A new microbial secondary metabolite, paulobutalipin (<b>1</b>), was isolated and characterized from the culture of a mountain soil-derived <i>Streptomyces</i> strain. The structure of paulobutalipin (<b>1</b>) was elucidated through a combined analysis of spectroscopic data, single-crystal X-ray diffraction, chemical modifications including application of the modified Mosher's method, and electronic circular dichroism calculations. In an in vitro hepatocellular steatosis model induced by palmitic and oleic acids, paulobutalipin (<b>1</b>) reduced intracellular lipid accumulation in AML12 hepatocytes by approximately 30% compared to that of vehicle controls. Moreover, it enhanced mitochondrial abundance in a dose-dependent manner, suggesting stimulation of mitochondrial β-oxidation. Our data identify paulobutalipin as a unique microbial natural product that promotes energy metabolism possessing structural complexity and minimal toxicity.